The mycobacteria are a diverse collection of acid-fast, gram-positive bacteria some of which cause important human and animal diseases [reviewed in Bloom et al., (1983), Rev. Infect. Dis., 5:765-780; and Chaparas, (1982), CRC Reviews in Microbiology, 9:139-197]. In man, the two most common mycobacteria-caused diseases are tuberculosis and leprosy, which result from infections with Mycobacterium tuberculosis and Mycobacterium leprae, respectively. These two diseases afflict more than 65 million individuals world-wide and result in over 4 million deaths annually, Bloom et al., (1983), Rev. Infect. Dis., 5:765-780.
The pathogenicity of these mycobacterial infections is closely tied to the host's immune response to the invading mycobacterium [Chaparas, (1982,), CRC Reviews in Microbiology, 9:139-197; Collins, (1982), Am. Rev. Respir, Dis., 125:42-49; Dannenberg, (1982), Am. Rev. Respit. Dis., 125:25-29; and Grange, (1984), Adv. Tuberc. Res., 21:1-78]. Not only does M. tuberculosis infect and grow within cells of the host's immune system, primarily the aveolar macrophage, but also it is the host's cellular immune response that plays the key roles in immunity from infection, containment of the infection at the initial focus of infection, progression or regression of the infection, and tissue damage or destruction at the foci of infection [Chaparas, (1982), CRC Reviews In Microbiology, 9:139-197; Collins, (1989,), Am. Rev. Respir. Dis., 125:42-49; Dannenberg, (1987), Am. Rev. Respit. Dis., 125:25-29; and Grange, (1984), Adv. Tuberc. Res., 21:1-78]. In addition, the standard method of detecting an M. tuberculosis infection, the tuberculin skin test, actually measures the host's cellular immune response to the mycobacterium [Snider, (1982), Am. Rev. Respir. Dis., 125:108-118]. The mycobacterial components that are important in eliciting the cellular immune response are not yet well defined.
A number of studies have attempted to define the mycobacterial antigens by standard biochemical and immunological techniques including the analysis of the target antigens of monoclonal hybridoma antibodies directed against mycobacteria [Daniel et al., (1978), Microbiol. Rev., 42:84-113; Engers et al., (1985), Infect. Immun., 48:603-605; Engers et al., (1986), Infect. Immun., 51:718-720; Grange, (1984), Adv. Tuberc. Res., 21:1-78; Ivanyi et al., (1985), Monoclonal Antibodies Against Bacteria (A. J. L. and E. C. Macario, eds.) Academic Press, Inc. New York. pp. 59-90; and Stanford, (1983), The Biology of the Mycobacteria (Ratledge and Stanford, eds.), Academic Press, London, vol. 9., pp. 85-127].
One particular antigen, a 65 kilodalton (KD) protein, is present in a wide range of mycobacterial species and has been most intensively studied as an antigen of M. leprae [Emmrich et al., (1986), J. Exp. Med., 163:1024-1029; Gillis et al., (1985), Infect. Immun., 49:371-377; Young et al., (1985), Nature, 316:450-459; and Mehra et al., (1986) Proc. Natl. Acad. Sci. USA, 83: 7013-7017]. This antigen has been designated the 65KD antigen or the cell wall protein-a (CWP-a) antigen since it appears to a co-purify with cell walls in some isolation procedures [Gillis et al., (1985), Infect. Immun., 49:371-377].
In Western blot assays, monoclonal antibodies directed against this antigen react with two major components in an M. leprae extract that migrate with apparent sizes of 55,000 and 65,000 daltons, and react occasionally with smaller components as well [Engers et al., (1985), Infect. Immun., 48:603-605 and Gillis et al., (1985), Infect. Immun., 37:172-178]. It is not known if these species represent discrete proteins or precursors and products, or result from chemical or enzymatic cleavage during isolation. In other species, such as M. gordonae, only a single species of about 65,000 daltons is detected with the monoclonal antibodies [Gillis et al., (1985), Infect. Immun., 49:371-377].
The 65KD antigen is one of the major immunoreactive proteins of the mycobacteria. This antigen contains epitopes that are unique to a given mycobacterial species as well as epitopes that are shared amongst various species of mycobacteria [Engers et al., (1985), Infect. Immun., 48:603-605 and Gillis et al., (1985), Infect. Immun., 49:371-377]. In addition, some other antigens that appear to be expressed by only one mycobacterial species are also found to contain epitopes expressed in other mycobacterial species. [Kingston et al., (1987) Infect. Immun., 55:3149.]
As discussed hereinafter, it is now found that purified 65KD antigen can elicit a strong delayed-type hypersensitivity reaction in experimental mammals infected with M. tuberculosis. Antibodies directed against this protein can also be detected in the sera of patients with tuberculosis or leprosy, and T-cells reactive with this antigen can be isolated from patients with leprosy or tuberculosis as well as from BCG-vaccinated persons [Emmrich et al., (1986), J. Exp. Med., 163:1024-1029; Engers et al., (1986), Infect. Immun., 51:718-720; Mustafa et al., (1986), Nature, 319:63-66; and Thole et al., (1985), Infect. Immun., 50:800-806]. Overall, the 65KD antigen appears to be a major, medically important B- and T-cell immunogen and antigen in humans.